LOCUS       sequence-368746-        6637 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of cell-surface protein extracellular domain 
            fused to Fc(DAPA)-Avi-6xHis. Protein is secreted from cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6637)
  AUTHORS   Wojtowicz WM, Vielmetter J, Fernandes RA, Siepe DH, Eastman CL, 
            Chisholm GB, Cox S, Klock H, Anderson PW, Rue SM, Miller JJ, Glaser 
            SM, Bragstad ML, Vance J, Lam AW, Lesley SA, Zinn K, Garcia KC
  TITLE     A Human IgSF Cell-Surface Interactome Reveals a Complex Network of 
            Protein-Protein Interactions.
  JOURNAL   Cell. 2020 Aug 20;182(4):1027-1043.e17. doi: 
            10.1016/j.cell.2020.07.025.
  PUBMED    32822567
REFERENCE   2  (bases 1 to 6637)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cell.2020.07"; journalName: "Cell"; date: "2020-08-20- 
            20"; volume: "182"; issue: "4"; pages: "1027-1043.e17"
COMMENT     Sequence Label: pD649-HAsp-TNF-Fc(DAPA)-AviTag-6xHis
FEATURES             Location/Qualifiers
     source          1..6637
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      41..176
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     103..122
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        238..285
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             304..903
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(304..323)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     800..820
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1033..1154
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1070..1089)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1124..1143
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(1207..1998,1999..2067))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1830..1849
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2068..2159)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2201..2220
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2226..2245
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2331..2919
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2820..2839
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3073..3090
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     3188..3205
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3189..3205
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3661..3677
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     enhancer        3844..4223
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4224..4427
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4378..4398
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4424..4448
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        4658..4676
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4658..4675
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     4689..4708
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             6056..6100
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             6101..6118
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6258..6275)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6264..6375
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6493..6513)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(6498..6637,1..190)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
ORIGIN
        1 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
       61 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
      121 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
      181 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc agcacgtgtt
      241 gacaattaat catcggcata gtatatcggc atagtataat acgacaaggt gaggaactaa
      301 accatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc
      361 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg
      421 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
      481 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
      541 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
      601 ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag
      661 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
      721 ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc
      781 ctggagacat ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc
      841 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
      901 tgacacgtgc tacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
      961 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     1021 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     1081 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     1141 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     1201 tggtcattac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     1261 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     1321 tggccccagc gctgcgatga taccgcgaga accacgctca ccggctccgg atttatcagc
     1381 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     1441 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     1501 gcgcaacgtt gttgccatcg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     1561 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     1621 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     1681 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     1741 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     1801 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     1861 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     1921 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     1981 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     2041 ggcgacacgg aaatgttgaa tactcatatt cttccttttt caatattatt gaagcattta
     2101 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     2161 aggggtcagt gttacaacca attaaccaat tctgaacatt atcgcgagcc catttatacc
     2221 tgaatatggc tcataacacc ccttgctcat gaccaaaatc ccttaacgtg agttacgcgc
     2281 gcgtcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     2341 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     2401 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     2461 cagataccaa atactgttct tctagtgtag ccgtagttag cccaccactt caagaactct
     2521 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2581 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2641 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2701 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2761 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2821 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2881 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2941 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     3001 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     3061 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag gcgagagtag ggaactgcca
     3121 ggcatcaaac taagcagaag gcccctgacg gatggccttt ttgcgtttct acaaactctt
     3181 tctgtgttgt aaaacgacgg ccagtcttaa gctcgggccc cctgggcggt tctgataacg
     3241 agtaatcgtt aatccgcaaa taacgtaaaa acccgcttcg gcgggttttt ttatgggggg
     3301 agtttaggga aagagcattt gtcagaatat ttaagggcgc ctgtcacttt gcttgatata
     3361 tgagaattat ttaaccttat aaatgagaaa aaagcaacgc actttaaata agatacgttg
     3421 ctttttcgat tgatgaacac ctataattaa actattcatc tattatttat gattttttgt
     3481 atatacaata tttctagttt gttaaagaga attaagaaaa taaatctcga aaataataaa
     3541 gggaaaatca gtttttgata tcaaaattat acatgtcaac gataatacaa aatataatac
     3601 aaactataag atgttatcag tatttattat catttagaat aaattttgtg tcgcccttaa
     3661 ttgtgagcgg ataacaatta cgagcttcat gcacagtggc gttgactcaa tattggccat
     3721 tagccatatt attcattggt tatatagcat aaatcaatat tggctattgg ccattgcata
     3781 cgttgtatct atatcataat atgtacattt atattggctc atgtccaata tgaccgccat
     3841 gttggcattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
     3901 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
     3961 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
     4021 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
     4081 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     4141 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     4201 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
     4261 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
     4321 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
     4381 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt ttagtgaacc
     4441 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga caccgggacc
     4501 gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt gccaagagtg
     4561 acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta gaacgcggct
     4621 acaattaata cataacctta tgtatcatac acatacgatt taggtgacac tatagaataa
     4681 catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc acctcggttc
     4741 tatcgaaaac gcgcctgcta gcgccaccat gaagaccatc attgccctga gctacatctt
     4801 ctgcctggtg ttcgctgcgg ccgccggacc tcagagagag gaatttccta gagatctgag
     4861 cctgatcagc cctctggctc aggccgtcag aagcagcagc agaacaccta gcgataagcc
     4921 cgtggctcac gtggtggcta atcctcaggc tgaaggacag ctgcagtggc tgaatagaag
     4981 ggccaacgct ctgctggcca acggcgtgga actgagagat aatcagctgg tggtgcccag
     5041 cgagggcctg tacctgatct atagccaggt gctgttcaaa ggccagggct gcccttctac
     5101 acacgtgctg ctgacccaca ccatcagcag aatcgccgtg tcctaccaga ccaaagtgaa
     5161 cctgctgagc gccatcaaga gcccctgtca gagagaaaca cctgagggcg ccgaagccaa
     5221 gccttggtac gaacctatct atctcggcgg cgtgttccag ctcgagaagg gcgatagact
     5281 gagcgccgag atcaacagac ccgactacct ggattttgcc gagagcggcc aggtgtactt
     5341 cggcatcatt gcccttggcg cgccgggcgg cgacaagacc cacacctgcc ccccctgccc
     5401 agctccagaa ctgctgggag ggccttccgt gttcctgttc ccccccaagc ccaaggacac
     5461 cctgatgatc agcaggaccc ccgaggtgac ctgcgtggtg gtggccgtgt cccacgagga
     5521 cccagaggtg aagttcaact ggtacgtgga cggcgtggag gtgcacaacg ccaagaccaa
     5581 gcccagagag gagcagtaca acagcaccta cagggtggtg tccgtgctga ccgtgctgca
     5641 ccaggactgg ctgaacggca aagaatacaa gtgcaaagtc tccaacaagg ccctggctgc
     5701 cccaatcgaa aagacaatca gcaaggccaa gggccagcca cgggagcccc aggtgtacac
     5761 cctgcccccc agccgggagg agatgaccaa gaaccaggtg tccctgacct gtctggtgaa
     5821 gggcttctac cccagcgata tcgccgtgga gtgggagagc aacggccagc ccgagaacaa
     5881 ctacaagacc acccccccag tgctggacag cgacggcagc ttcttcctgt acagcaagct
     5941 gaccgtggac aagtccaggt ggcagcaggg caacgtgttc agctgcagcg tgatgcacga
     6001 ggccctgcac aaccactaca cccagaagtc cctgagcctg agccccggcg gcagcggtct
     6061 taatgatatt ttcgaggctc agaagattga gtggcatgaa caccaccacc atcaccacta
     6121 aggccggcct gtagtcactg gcacaacgcg ttacccggta agccaatcgg gtatacacgg
     6181 tcgtcatact gcagacaggg ttcttctact ttgcaagata gtcttgagta gtaaaataaa
     6241 tagatagaga aaatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
     6301 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     6361 tgaggaaatt gcatcacaac actcaaccct atctcggtct attcttttga tttataaggg
     6421 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     6481 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     6541 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     6601 gctccccagc aggcagaagt atgcaaagca tgcatct
//
